Flexible Method Development Using a New pH Stable HPLC Column, Kromasil Eternity C18

By: Jared Benedict, Marketing, Sales Manager, Kromasil, Reporter US Volume 28.1

Jared Benedict, Marketing and Sales Manager, Kromasil



The separation of new generation pharmaceutical compounds has increasingly become more challenging. There is a need for stationary phases that can withstand a wide pH range, from very acidic to very basic conditions, while maintaining peak shape, symmetry and lifetime. Utilizing pH as a method development tool has now allowed more flexibility in altering retention times of ionizable compounds. More basic compounds are being synthesized during the discovery of pharmaceuticals and thus there is a great need for robust stationary phases when running at high pH. This article shows how Kromasil Eternity provides solutions for the separation of ionizable compounds using extreme pH conditions, from pH 1 to pH 12.

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In this study, we show the retention time of amines and acids can be altered by changing the pH, utilizing a single column (Figure 1). Running under basic (pH 11.0) conditions will destroy classic C18 phases very quickly, thus limiting the range of pH that can be utilized. However, when running basic pH (11.0) conditions on a Kromasil Eternity C18, procaine, an amine, is strongly retained whereas the acidic nitrobenzoic acid is weakly retained. The reverse is true under acidic conditions. Therefore, not only can you utilize the solvent strength and temperature as a method development tool but you can now also utilize nearly the full pH range. The most striking conclusion from Figure 1 is that these extreme pH conditions can be run on a single column. Kromasil Eternity can be used at an extended pH range while maintaining a very long lifetime.

Figure 1. Separation of Ionizable Compounds at Three Different pH Conditions

In addition to utilizing the flexibility in pH in analytical method development one can also employ the robustness of these columns for semi-preparative HPLC. When doing high throughput purification of pharmaceutical compounds one needs a column that can run under extreme pH conditions to maximize the separation factor and thus the loadability. Kromasil Eternity C18 provides a solution for high loadability and flexibility under extreme pH conditions using a single column which also maintains a long lifetime that gives the maximum total economy for your purification.

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Kromasil Eternity provides flexible method development and long column lifetime through the use of unique modified silica. Kromasil Eternity C18 is available as a 2.5 μm particle size for UHPLC or HPLC applications. The small particle size allows for fast runs that maximize efficiency and minimize solvent consumption. Additionally, a 5 μm particle size is available for analytical to semi-preparative HPLC up to 30 mm I.D. columns. The loadability and lifetime of the semi-preparative columns are excellent.

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